Type 1 diabetes (T1D) is an autoimmune disease characterized by T cell-mediated destruction of the pancreatic islet beta cells. Our knowledge of at least some of the beta cell antigens targeted by T cells in both the NOD mouse model and T1D patients can now be used practically to develop antigen-based strategies to interfere with pathogenic T cell populations and augment natural tolerance induction pathways. We hypothesize that an effective immunotherapeutic strategy for T1D would be one that eliminates pathogenic CD8+ and CD4+ T cells specific for an important beta cell antigen, while at the same time fostering the expansion of regulatory T cells (Treg) capable of controlling the remaining pathogenic T cells that target a variety of beta cell antigens. We will develop such an approach by taking advantage of our experience in the targeting of antigens to dendritic cells (DC) using antigen-linked antibodies to the DC endocytic receptor DEC- 205. When antigens are delivered in this way in the absence of an adjuvant, DC present them in a tolerogenic manner and cause naive T cells to be deleted, rendered unresponsive, or endowed with regulatory characteristics. Using a reagent that we have recently produced, we propose to deliver the beta cell antigen proinsulin to DC and determine the effect of this treatment on pathogenic CD8+ and CD4+ T cells and on the induction of Treg. These studies will employ an NOD-based mouse model that we have developed (designated NOD.22mnull.HHD.Ins2+/-) that transgenically expresses T1D-associated human HLA-A*0201 and is also heterozygous for an Ins2 knockout allele which leads to diminished thymic insulin expression, hence mimicking the situation in the majority of T1D patients. State-of-the-art mouse models incorporating human cells will also be employed. Finally, our experimental systems will be ideal to determine the pathways by which targeting of antigens to DC can lead to induction of T cell tolerance. Three Specific Aims are proposed: (1) To target proinsulin to DC via DEC-205 and monitor its effects on T cells and the development of T1D in NOD.22mnull.HHD.Ins2+/- mice; (2) To determine whether human islet-reactive T cells can be tolerized in response to DEC-205-mediated delivery of beta cell antigens to DC; (3) To determine the pathways important for the T cell tolerance observed in response to DEC-205-mediated delivery of beta cell antigens to DC. Our proposed studies will have important implications for the development of antigen-specific therapeutics for T1D and will provide information that will help to guide the future development of immunomodulatory therapies for this disease.